Lymph node responses to malignant homo- and heterografts in the hamster.

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Lymph Node Responses to Malignant Homo- and
Heterografts in the Hamster’
DAVID SHEPRO: LESTER P. EIDELHOCH” AND DONALD I. PATT
Department o f Biology, Boston University, a n d Sears Surgical Research
Laboratory o f the F i f t h Surgical Service, Boston City Hospital
The Syrian hamster, Mesocricetus auratus, is used extensively for transplantation
research because of the ability of the noninbred animal to tolerate homografts occasionally and heterografts of normal and
malignant tissues (Lutz et al., ’50; Toolan,
’51; Adams et al., ’56; Foley and Handler,
’57; Billingham arid Hildemann, ’58). The
remarkably high incidence of successful
transplants in the hamster cheek pouch
has led some investigators to theorize that
the cheek pouch might be an immunologically privileged area similar to the anterior
chamber of the eye. Adams and his coworkers (’56) explained successful orthotopic homografts of skin (10-12 months
duration) by postulating a possible “defect” in the hamster’s ability to respond to
tissue homografts. In contrast, Billingham
and Hildemann (’58) in a similar experiment found that intercolony homografts
did not persist in the hamster; that the
hamster rejects a skin graft in a manner
similar to that which has been described
for other mammals. However, intracolony
homografts did persist until the close of
the experiment and the authors explained
this phenomenon by stating that the hamster “apparently has a paucity of transplantation antigens.”
Notwithstanding differences in interpretation, it is generally agreed that the Syrian hamster is distinctive among mammals in that it readily accepts a variety of
transplanted tissues. In light of this evidence it was decided to investigate the
lymphoid response of the hamster to tumor
homo- and heterotransplants and to orthotopic skin homografts; the latter portion of
the experiment is still in progress and will
be reported on at a later date. In addition
it was anticipated that the data might provide information pertinent to first, the feasibility of using the histology of a regional
node draining a human heterotransplant
as a base line for anti-tumor studies and
second, the identification and localization
of lymphatic cell(s) involved in the hamster’s immunological mechanism.
MATERIAL AND METHODS
Animals. Golden hamsters (Mesocricetus auratus) of both sexes, about 5 weeks
old between 50-70 gm, were used. The
hamsters were obtained from one local
professional breeder. The animals were
housed two to a cage and were fed Purina
Laboratory Chow and water ad libitum.
Implantation and conditioning. The
technic of transplanting the tissues to the
pouch was essentially the same as described by Lutz et al. (’51). The tissues
were cut into fragments 1 to 2 mm and
were deposited near the tip of the everted
pouch with a no. 18x trochar. Only the
left cheek pouch received a graft, thus the
right side of the animal served as an additional control. Those animals receiving
a heterotransplant were conditioned with
cortisone acetate according to the following scale: 60 to 70 gm animal weight, 2.0
mg of cortisone; 70 to 80 gm, 2.5 mg of
cortisone; 80 to 90 gm, 3.0 mg of cortisone.
The initial injection was given on the day
of the operation and thereafter every three
days until the experiment terminated. All
injections were subcutaneous, as close to
the mid-dorsal line as possible.
Tissues. The tumors used were a human
amelanotic melanoma (BCH 2 ) and a
spontaneous hamster amelanotic melanoma. Both tumors have an accelerated
growth rate for the first 10 to 14 days and
__ __
’This investigation supported in part by
USPHS grant C-l644(M & G ) .
Present address : Simmons College, 300 The
Fenway, Boston, Mass.
Present address: Bronx Municipal Hospital
Center, Bronx 61, New York City.
393
394
DAVID SHEPRO AND OTHERS
then level off and grow in a plateau fashion. Generally by the end of the third week
the heterologous tumor will have increased
from a 1-mm fragment to a firm, pearly
white nodule of about 18 mm. The homologous tumor at this point will measure
about 1 cm. From the third week on, the
tumors either enlarge, become edematous,
or ulcerate.
Autopsy. Following transplantation the
animals were killed at predetermined intervals. Two animals were used for each
stage in the series. The following code
was devised to identify the animals:
Sp, transplanted inert cellulose sterile
sponge.
C, cortisone control.
HT, hamster amelanoma melanoma
transplanted into the left cheek pouch.
T , human amelanoma melanoma transplanted into the left cheek pouch.
TC, human amelanoma melanoma
transplanted into the left cheek pouch of
cortisone conditioned hamsters.
The above code letters were placed in
the numerator of a fraction and the killing
date in the denominator. When cortisone
conditioning was employed, the number
of cortisone injections a t the time of killing was also placed in the numerator along
with the code letters. Thus, a n animal
labeled “TC318” would signify a n animal
with a tumor heterograft plus three injections of cortisone acetate and was killed o n
the 8th day post-operative. Two animals
were used on each killing date.
The superficial cervical nodes, the deep
cervical nodes, the axillary nodes, the auricular nodes and the spleen were removed
and wet weights recorded. The superficial
cervical nodes were found to be the principal draining nodes of the cheek pouch;
the other tissues removed served as additional controls. The tissues were fixed in
Carnoy’s solution, sectioned at 6 11 and
stained with methyl-green-pyronine. As a
rule only the regional and non-regional
superficial cervical nodes were studied in
detail except where conditions warranted
checking the additional control tissues.
The condition, size and weight of the tumors were recorded and then the tumor plus
the adjacent tissue were removed, fixed
in Zenkers and stained routinely with hematoxylin and eosin.
Histological examination. The regional
lymph node and the con tralateral node
were analyzed for the following characteristics :
1. The number of secondary centers.
2. The per cent ratio of large lymphocytes to small lymphocytes in the cortex.
These determinations were made by counting all the large cells within the boundaries
of a 1 m m reticule (divided into 100
squares) and then counting the small
lymphocytes in 10 of the smaller units and
multiplying by 10. Ten such counts were
made from various areas of the cortex,
selected at random, and the results recorded as their mean value.
3 . Differential counts were made of the
medulla to determine the percentages of
( a ) small, medium and large lymphocytes,
( b ) plasma cells, ( c ) reticular cells, ( d )
granulocytes and ( e ) mitotic cells. However, in the accompanying tables only the
data computed for the large lymphocytes
and plasma cells are recorlded.
In the differential courlts only those
cells with a “cartwheel” nucleus and a perinuclear clear area separating the nucleus
from the mass of cytoplasim were called
plasma cells. The cells which we have designed as large lymphocytes are similar
to those which other investigators have
called “immature lymphocyte,” “young
lymphocyte,” “preplasma cell,” “immature
plasma cell,” “transitional cell,” etc. The
large lymphocyte of the hamster is between 9-12.5 ~r in diameter, has one or
two distinct nucleoli, and the nucleus is
s u r r o u d e d by a broad layer of pyronine
positivc cytoplasm. Except for size, it appears to be identical to the “large lympho i d cell described by Scothorne and McGregor (’55) who coined the latter term in
preference to those listed above because
it was “non-committal.”
Normal gross and microscopic ancitorny
The normal superficial cervical nodes
are pink or pearl gray with a mottled gray
surface at the hilum. They measure about
1 cm in length and the mean weights of
the left and right superficial cervical nodes
are 31.9 m g and 31.2 me, respectively.
The weight range was betv,een 19.6 and
57.4 mg.
LYMPH NODE RESPONSES T O TUMORS
The structural plan of the node is essentially the same as that which has been
described for other mammals; therefore,
only the briefest resume is submitted.
Cortex. Forming the outer limits of the
cortex is a fibrous capsule which sends off
relatively few trabeculae. Beneath the capsule, but not always visible, is the marginal
sinus containing small and medium lymphocytes, granulocytes, and reticular cells.
The cortex proper is composed of primarily
small lymphocytes; i n addition there are
scattered reticular cells, and a n extremely small number of large lymphocytes
(1.9% ) . Even this small percentage is
somewhat deceiving for there are broad
areas of the cortex that are literally devoid
of any large lymphocytes. There is a tendency for the large lymphocytes to congregate close to and between the secondary
centers and our counts were made from
fields selected randomly from these areas.
Scattered throughout the cortex but usually close to the periphery are the secandary centers. In some nodes there are as
few as three and in other nodes there are
as many as 19 centers. The average is 8.8
centers per node. They ranged in size between 50-375 in diameter and, utilizing
the terminology of Maximov and Bloom
(’52), most of the centers are in a mildly
active state.
Medulla. The results of the differential
counts, in descending order of occurrence,
are : small and medium lymphocytes,
61.7% ; plasma cells, 17.0% ; reticular
cells, 8.6% ; large lymphocytes, 6.7% ;
granulocytes, 4.5% ; mitotic cells, 1.5%.
The cords have a reddish cast because of
the large number of cells with pyronine
staining cytoplasm. The sinuses vary in
diameter and normally contain relatively
few cells, e.g., small lymphocytes, reticular
cells and granulocytes. Mature plasma
cells were never observed in the medullary
sinuses.
RESULTS
Response t o cortisone conditioning
Following 9 injections of cortisone acetate given over a period of 29 days (table
I), the cervical nodes (60% ) and the
spleen (38% ) were reduced markedly in
weight (fig. 1). The size of the cortex
was reduced continually with a corresponding increase i n the medulla. At the
395
Fig. 1 The superior cervical nodes and spleen
of a normal animal compared with similar tissues
taken from a n animal that had received 8 injections of cortisone acetate over a period of 28 days.
In addition to the extreme reduction in size, the
experimental nodes were extreinely hard and
dark.
termination of the experiment the nodes
were practically all medullary. Simultaneously the number of secondary centers
was reduced and the few remaining in
each node were of the “waning” or “terminal” variety. For example, in animals
”(2318, an average of 7 centers was reported but of the 7 only two could be considered even slightly active.
The cellular population shifted with all
cells declining in numbers except the granulocytes and plasmacytes. There was a n
initial plasmacytosis for the first two days
followed by a gradual decline. It was
quite evident that the cortisone exerted an
inhibitory effect on the plasma cells for
following each injection of the hormone
the plasma cells decreased and during the
time between injections (two or three
days) the cells increased in number. However, the initial plasmacytosis of the first
two days was sufficient to raise the mean
value equal to the normal number of
plasma cells.
After the first week of treatment, a
steady rise in the number of granulocytes
culminating in a 10-fold increase at the
termination of the experiment was observed. Quantitatively, second in importance to the granulocytic change was the
77
74
50
72
80
67
58
C7/22
C8/28
C9/29
71.5
51
X.
140
68
82
C4/14
75
63
66
C4/10
65
62
64
C3/8
C6/21
62
50
C2/7
C6/18
66
42
61
84
63
98
C1/3
C2/6
87
c1/2
90
Spleen
C2/4
88
gm
Body
wt.
c1/1
Exp.
and day
5
4
1
13.7
13.1
12.1
23.9
17.4
13.1
13.9
5.9
9.2
12.3
8.6
8.2
6.3
6.0
4.4
18.5
1.o
1
1.1
4.0
0.8
0.5
6
5.1
8.0
3.0
2.5
12.0
14.0
18.5
15.5
20.0
21.0
16.5
16.0
20.0
8.0
29.0
%
31.0
Plasmacytes
1.2
5
4.5
4.0
1.o
7.4
1.2
4
8.2
5.7
10.7
5
5.5
1.0
5
9.7
11.1
3.0
5.0
2.5
12.7
1.4
1.8
1.3
3.0
6.0
12.0
1.o
1.6
1.3
6.0
0.6
3.5
%
%
0.6
Large
lymphocytes
Large/
small
lymphocytes
Superficial cervical nodes
16.3
5
7
11
18.4
19.4
11.7
5
16.2
17.1
20.6
7
Secondary
centers
20.3
Right
24.5
Left
-
Cortex
Response of hamsters to cortisone conditioning
TABLE 1
10.2
12.0
10.0
9.5
9.5
8.5
6.5
13.0
12.0
14.5
12.0
9.5
7.0
5.0
14.0
%
Reticular
cells
Medulla
1.3
0.0
2.0
0.5
1.0
1.0
1.5
2.0
1.0
1.0
2.0
2.0
2.0
1.5
%
0.5
20.7
51.0
38.0
34.5
30.5
35.0
10.0
31.0
5.0
11.0
14.5
5.0
10.0
10.0
%
3.5
Mitoses Granulocytes
v1
0
~
0
k-
3
0
$
z
ti
6,
U
k
LYMPH NODE RESPONSES TO TUMORS
marked depletion of cortical large lymphocytes (50% ).
Response to implanted cellulose sponge
The overall response of the hamsters to
the grafting of an inert sponge in the left
cheek pouch was remarkably slight (table
2). Even the mild hyperplasia of the cervical nodes and spleen are minimized by
the realization that the mean weights lie
within the range of the normal mean
weights. In addition, the non-regional
nodes outweighed the regional nodes, minimizing the slight difference.
Except for the increase in secondary
centers, both the cortex and the medulla
appeared normal. There was not any increase in pyronine accumulation, differential counts were normal and sinuses were
not dilated and contained very few cells.
The cheek pouches at the site of implantation displayed a slight degree of inflammation for the first three days. After
397
this period even this mild, localized reaction disappeared.
Response to an homologous tumor
The homotransplantation of malignant
tissues evoked an unequivocal response in
the draining nodes (table 3 ) . Moreover, in
three important categories, the weight of
the nodes, the number of secondary centers and the per cent of cortical large
lymphocytes, the means computed were almost twice as great as those obtained from
the contralateral nodes. Specific animals
in the series reflected even greater differences. A comparison of the cortical large
lymphocyte response in hamsters "HT/28
shows that the ipsilateral nodes responded
9 times more than the contralateral nodes.
The degree of reaction of the regional
node to the tumor transplant is more dramatically illustrated when compared with
the normal statistics and the observed in
terms of per cent increases. The mean
TABLE 2
Response of hamsters to a n inert cellulose sponge transplanted into the l e f t cheek pouch
Weight
Exp.
and day
Body wt.
Cervical nodes
Left
Sp/l
sp/2
sp/3
Sp/6
SP/7
Sp/8
Sp/lO
Sp/14
Spleen
Right
g m
mg
mg
mg
80
71
80
78
86
82
93
80
50.0
36.0
27.3
38.6
25.0
31.0
40.0
30.5
34.8
46.0
37.8
26.3
34.6
30.6
48.0
45.6
31.8
37.5
124
122
156
134
128
170
109
101
130
x
Superficial cervical nodes
Exp.
and day
Sp/l
SP/2
sp/3
Sp/6
sp/7
Sp/8
sp/m
Sp/14
~X
Cortex
Secondary
centers
Left
Right
37
17
10
13
13
12
9
10
15.0
14
18
14
12
12
14
14
11
13.6
Medulla
Large/small
lymphocytes
Left
Right
Large
lymphocytes
Left
Right
Plasmacytes
Left
Right
%
%
%
%
%
%
1.9
1.7
1.0
1.0
2.6
1.8
1.4
1.3
1.6
2.0
2.8
1.4
1.2
1.6
0.8
1.4
1.2
1.6
8.5
11.0
8.0
10.5
8.5
7.0
6.5
7.5
8.0
8.3
23.5
25.0
18.0
13.5
18.5
13.0
14.0
14.0
17.4
21.5
37.0
11.0
11.5
7.5
15.5
10.5
15.0
16.2
9.0
9.0
9.5
8.0
12.0
6.5
8.5
8.8
398
DAVID SHEPRO AND OTHERS
TABLE 3
Response of hamsters to an homologous tumor (amelanoma melanoma) transplanted
into the left cheek pouch
Weight
Exp.
and day
Body
wt.
Sex
Cervical nodes
Left
HT/1
HT/6
HT/8
HT/10
HT/12
HT/14
HT/28
HT/32
HT/34
HT/38
HT/42
0
0
d
0
0
d
0
d
0
d
d
Right
gm
mg
mg
mg
76
80
77
81
80
81
43.9
36.0
29.0
56.4
45.8
43.2
50.4
25.0
30.2
25.2
36.0
36.6
100
158
131
138
148
166
105
94
86
113
105
97.4
69.6
52.0
156.0
70.0
33.8
38.4
23.8
60.0
48.2
234
259
185
494
190
63.0
37.9
200
X
Remarks
SDleen
Just a spot in the pouch
Tumor negligilde; opaque area
Tumor: 10mm
Tumor: 3 masses, each 10 mm
Tumor: 3 masses, each 10 mm
Tumor: 2 masses, each 20 mm
one mass, 30 mm
Tumor: 30 mm, ulcerating
Tumor: 30 mnn, ulcerating
Tumor: 25 mnn, ulcerating
Tumor: 45 mnn, ulcerating
Tumor: 10 mnn, ulcerating
30 mm, ulcerating
______
Superficial cervical nodes
EX~.
and day
Cortex
Secondary
centers
Medulla
Large/small
lymphocytes
Large
lymphocytes
___IPlasmacytes
~~~
Left
HT/ 1
HT/6
HT/8
HT/lO
HT/12
HT/14
HT/28
HT/32
HT/34
HT/38
HT/42
X
32
39
38
22
23
23
26
23
20
35
32
28.5
Right
39
3
19
22
21
17
20
24
15
14
24
19.8
Left
Right
Left
Right
%
%
%
%
2.5
4.2
2.8
5.3
3.5
3.4
9.2
4.5
3.9
6.6
6.2
4.7
3.8
1.5
3.3
4.7
3.0
2.2
0.9
1.7
4.1
3.5
4.1
2.9
5.0
10.0
8.5
2.5
5.0
5.0
4.0
11.0
9.5
5.5
5.5
6.5
4.0
10.0
4.0
3.5
6.5
11.0
11.5
9.5
6.0
7.0
7.5
7.3
weight of the regional nodes increased
227%, the number of cortical large lymphocytes increased 147%, and the plasma
cells increased 115%. Although the medullary large lymphocyte did not vary in
number, a definite increase in the number of medium sized lymphocytes with pyronine staining cytoplasm occurred.
One could summarize the changes that
occurred by stating that pyronine positive
cells in both the cortex and the medulla
increased and that the most significant alteration of the cell population was in the
massive accumulation of large lymphocytes adjacent to and between the numerous secondary centers of the cortex.
The signs of reactivity of the contralateral nodes and the spleen suggested that
Left
5%
18
28
23
31
44
57
35
42
46
44
44
37.5
Right
%
13
20
20
31
32
41
19
39
55
35
45
31.8
the tumor antigen( s ) were systemic and
also the possibility of cross lymphatic
drainage of the right and left cheek
pouches.
Neither the deep cervical, axillary nor
auricular nodes showed signs of responding to the tumor.
Response to human tumors
The major changes from the normal
were the hyperplasia and the increased
number of secondary centeics (table 4).
The weights of the regional nodes, contralateral nodes and spleen increased about
5 0 % , 33% and 43% , respectively. The
per cent increase of the secondary centers
was almost 200% on the operated side and
almost 100% on the non-operated side.
399
LYMPH NODE RESPONSES TO TUMORS
TABLE 4
Response o f hamsters to a human tumor (Amelanoma melanoma) transplanted
into the l e f t cheek pouch
Weight
Exp.
and day
Body
Sex
Cemcal nodes
Wt.
Left
T/1
T/2
T/3
T/6
T/ 7
T/8
T/fJ
T/10
T/12
T/14
T/21
T/22
-
d
d
d
d
d
d
?
0
0
0
0
0
g m
mg
m g
mg
76
75
83
90
89
89
85
91
70
80
80
83
32.4
39.4
36.6
41.4
59.2
53.0
51.0
50.0
52.4
50.2
40.2
46.2
46.0
24.0
36.6
41.4
39.2
42.2
42.2
41.2
46.2
46.8
52.0
42.0
39.0
41.1
109
111
123
140
167
132
170
179
147
199
168
180
152
X
Remarks
Spleen
Right
No growth
No growth
No growth
Mosaic'
Mosaic'
Mosaic'
No growth
No growth
No growth
No growth
No growth
No growth
Supefficial cervical nodes
Exp.
and day
Left
T/ 1
T/2
T/3
T/6
T/7
T/8
T/9
T/10
T/ 12
T/14
T/21
T/22
-.
X
Cortex
Secondary
centers
27
27
31
27
20
58
21
19
21
25
28
10
26.2
Medulla
Large/ small
lymphocytes
Right
Left
%
%
14
16
18
11
10
22
21
10
22
20
19
9
16.2
3.5
1.7
3.3
1.1
2.7
3.0
1.7
1.3
1.1
1.1
1.4
0.9
1.8
2.9
2.2
3.7
1.3
1.1
1.9
1.6
2.5
1.5
1.3
1.1
0.6
1.8
Right
Large
lymphocytes
Left
Right
Plasmacytes
Left
%
%
%
13
13
18
11
9
11
10
15
10
7
8
5
10.1
11
17
13
15
6
6
10
6
16
4
5
10
9.9
21
32
25
37
27
33
16
17
27
26
19
8
24.0
Right
%
25
25
21
24
11
30
18
21
22
16
19
13
20.3
Tumor cells, WBC, RBC.
However, a parallel increase of large
lymphocytes in the cortical zone did not
occur. The mean percentage ratio of
large: small lymphocytes was computed at
1.8%, essentially normal. Only during
the first week did any hamsters show signs
of cortical lymphopoiesis which might have
represented an early response to the graft.
Although the differential counts of the
medullary cells were only slightly above
normal, the general appearance of the
medulla was not normal. An increase of
pyronine accumulations primarily due to
the deep staining of medium sized lymphocytes was the important deviation. In
addition, the cords were enlarged and the
sinuses were packed with cells. In con-
trast, the other nodes were not affected in
this manner.
Histological examination revealed that
none of the human tumors grew. Implantation sites removed from the hamsters for
the first three days had a few isolated
clumps of tumor cells. Similar slides made
from animals killed between days 4-8 were
characterized by a mosaic of blood cells,
connective tissue elements and few isolated tumor cells. All other slides were
negative.
Response to h u m a n tumor plus
cortisone conditioning
With one exception, the grafting of human amelanoma melanoma to a cortisone
400
DAVID SHEPRO A N D OTHERS
TABLE 5
Response of hamsters to a human tumor (amelanoma melanoma) transplanted into the
left cheek pouch coincidental with cortisone conditioning
~
_
Weight
Exp.
and day
Body
wt.
Remarks,
Cervical nodes
Left
Spleen
Right
gm
mg
mg
mg
TC1/1
TC1/2
TC1/3
TC2/6
TC2/7
TC3/8
75
74
76
81
70
71
22.5
36.7
12.5
14.9
36.6
20.8
22.8
29.0
22.4
24.4
39.0
25.8
81
61
54
88
99
74
TC4/10
74
16.9
14.5
50
TC4/14
TC6/18
TC6/21
TC7/22
TC8/28
.-
77
80
75
69
77
18.0
7.6
7.3
12.5
7.0
17.8
19.4
14.2
5.9
30.0
13.5
21.7
84
49
58
X
No growth
No growth
No growth
No growth
No growth
Tumor: minute opaque
disc
Residual tissue only;
opaque disc
Tumor: 22 :mm
Tumor: 25 imm
Tumor: 25 iinm
Tumor: 45 mm
Tumor: 80 mm
81
53
69
Superficial cervical nodes
Cortex
Exp.
and day
TC1/1
TC1/2
TC1/3
TC2/6
TC3/8
TC4/10
TC4/14
TC6/18
TC6/21
TC7/22
TC8/28
__
X
Medulla
_
I
_
_
Secondar,
centers
Large/small
lymphocytes
_____
Left
Right
Left
%
%
%
4
3
19
7
3
3
2
1
1
6
1
18
13
7
4
1
1
1
2
5
5.4
0.6
1.0
1.3
0.9
1.2
1.1
0.8
0.7
0.8
2.0
0.8
1.1
2.7
0.6
2.0
1.6
2.8
1.1
0.6
0.2
0.8
1.2
1.6
1.3
6.5
3.0
2.5
2.0
4.5
6.0
5.0
3.0
2.5
1.5
5.0
3.8
2
2
4.3
Right
Large
lymphocytes
conditioned hamster did not evoke any
lymphoid response that could not be accounted for by the hormone alone (table
5). The exception was the plasma cell
mean which was 6% less than the mean
computed for the cortisone control. The
plasmacytosis observed for the first two
days in the control was not evident in the
experiment where cortisone was used as a
conditioning agent.
Tumor transplants were 100% successful. By the end of the second week the
tumors measured about 25 mm; at the
end of the third week about 40 mm; at
the end of the 4th week and the experiment about 80 mm.
Left
I’lasmacytes
Right
Left
‘To
%
6.0
2.0
3.5
10.0
9.0
6.5
8.0
2.5
2.0
2.5
11.0
5.7
101.5
10’.0
9.5
27.5
5.0
4.5
6.5
13.0
15.5
18.0
13.0
12.1
Right
%
11.0
12.0
7.5
8.0
2.0
5.0
6.0
6.5
13.5
12.5
9.5
8.5
DISCUSSION
From the data presented on the normal
histology of the hamster’s lymphatic tissue
the lymph nodes show no indication that
they are unique. The structural framework of the nodes and the morphology and
per cent of the cells follows a pattern similar to that described for other common
laboratory animals. It is important to
emphasize that in the control nodes the
number of large lymphocytes of the cortex
is exceedingly small. In fact there are
large areas of diffuse cortical tissue where
the large lymphocyte is so rare that the
ratio of 1arge:small lymphocytes is less
than 0.05%.
_
LYMPH NODE RESPONSES TO TUMORS
The results reported for the cortisone
control are consistent with the literature
indicating adrenal cortex control over
lymphatic elements (Dougherty, '45, '53;
Scothorne, '56; Teir, '54). The extreme
atrophy of the nodes, the depletion
of secondary centers and the extreme reduction in the number of cortical large
lymphocytes are examples of the hormonal
control (table 6). Even though the plasma
cell mean is at a normal level (because of
the unaccountable initial increase in
plasma cells) the overall effect of the hormone was of an inhibitory nature. Because the effects of cortisone on the plasma
cell series is not as clearly defined and thus
not as predictable as is stated to be the
case for other lymphoid elements we hesitate simply to disregard this initial plasmacytosis. Whereas Teilum et al. ('50)
reported marked regression of the massive
accumulations of plasma cells in rabbit
spleens, Craig ('52) concluded that adrenocortical activity (cortisone and ACTH)
did not inhibit the formation of the plasma
cell series in rabbit lymph nodes.
The gross and histological changes of
the regional nodes draining the sites of the
implanted homologous amelanomas is similar to the lymphoid response to known antigens and other homotransplants as reported by Andreini et al. ('55), Craigmyle
('58), Ringertz and Adamson ('50), Scothorne ('56) and Scothorne and McGregor
('55). Because the non-regional nodes
and the spleens were also affected, but to
a far lesser degree, it is assumed that the
tumor substances that stimulated the regional nodes were not entirely confined
to the operated side. The unique aspect
of the animal's response to the tumors is
that none of the nodes in the area of the
cheek pouch, e.g., axillary and auricular,
increased in weight or showed signs of reactivity.
The major changes from the normal included an increase in secondary centers,
cortical large lymphocytes and medullary
plasma cells (table 6). That the nodes
are actively engaged in protein metabolism
is evident by the large accumulations of
pyronine positive ceIls throughout the node.
On the basis of these data it is tentatively
concluded that the ability of the hamster
to accept a homograft readily is not due to
Y
!
Y
z
k
40 1
402
DAVID SHEPRO AND OTHERS
any “defect” in the immunological response
of the lymphatic tissues. Our results further concur with the many investigations
which have reported the importance of the
large lymphocyte and plasma cell in the
process of immunity. Moreover, we agree
with Scothorne that greater attention
should be focused on the “large lymphoid
cell” of the cortex as a possible antibody
producer because the greatest and most
impressive variations of the lymph node
cell population to the transplants and to
cortisone were seen in the large lymphocyte count in the cortex.
It is more difficult to evaluate properly
the results of the human tumor study in
non-conditioned hosts. Based upon gross
weights and the number of secondary centers the nodes appear to respond to the
tumor per se (table 6) for these means
were significantly higher for the regional
nodes than for the non-regional and control
nodes. In contrast, the regional node plasma cell mean was only slightly higher than
in the non-regional nodes and the percentage of 1arge:small lymphocytes was normal for both sides. However, as was stated
previously (table 4 ) viable tumor cells
were evident only during the first 8 days.
If the means are calculated for the period
when supposedly antigenic stimulation was
present then the inconsistency disappears.
For now, in addition to the significant
changes mentioned above, appreciable increases in the important cortical large
lymphocytes and plasma cells are present.
On the operated side the large lymphocyte
mean increases 40% and the plasma cell
mean increases 70%. On the non-operated side the increases are only 10% for
the large lymphocyte and 28% for the
plasma cells. In some earlier experiments
not reported here, transplanted human and
hamster muscle tissues evoked no abnormal variation to the draining nodes.
The experimental nodes taken from
hamsters receiving malignant heterotransplants plus cortisone were especially interesting in that the data derived from the
experimints were essentially the same as
those which had been calculated in the cortisonecontrol experiment. me only exception was that the plasma cell mean was
lower than observed in the control (this
discrepancy discussed previously). The
results indicated quite conclusively that
cortisone inhibits any antigenic effects of
the growing transplant on the regional
nodes. Thus the histology of a regional
lymph node draining a tumor heterotransplant is an undesirable baseline for antitumor studies when the host is conditioned
with cortisone.
Our results agree with Sclothorne (’56)
that the hormone decreases the number
of large lymphocytes or “large lymphoid
cells” of the cortex. Our experiments were
not designed to test whether the cortisone
acted primarily by lowering the antigenicity of the graft, by direct action on the
node or a combination of both.
SUMMARY
1. The normal histology of lymphoid tissues and the weight changes and cytological variations of the superlicial cervical
nodes draining sites of homo- and heterologous tumor transplants were studied in
the hamster.
2. The histology of the hamster lymph
nodes is similar, quantitatively and qualitatively, to comparable tissue in the mouse,
rat, guinea pig and rabbit.
3. Cortisone conditioning inhibits any
alterations of the regional node that might
be induced by a successfully growing tumor heterotransplant.
4. Tumor heterotransplants in non-conditioned hamsters evoke a mild response in
the regional nodes for 8 days characterized
by increased weight of the node and slight
increases in the number of secondary centers, large lymphocytes and plasma cells.
5. Following tumor homotransplantation, the regional node responds in a fashion that may be called a typical histo-immunological response. Thus it is concluded that there is no defect in the ability
of the hamster’s lymphatic tissue to respond to homotransplantation stimulation.
6. It is suggested that the cortical large
lymphocyte as well as the plasma cell is
involved in the immunological response.
LITERATURE CITED
Adams, R. A., D. I. Patt and B. R. Lutz 1956
Long term persistence of skin hcmografts in
untreated hamsters. Transplant. Bull.. 3; 4142.
LYMPH NODE RESPONSES TO TUMORS
Andreini, P., M. L. Drosher and N. Mitchison
1955 Studies on the immunological responses
to tumor transplants in the mouse. J. Exp.
Med., 102: 199-204.
Billingham, R. E., and W. H. Hildemann 1958
Studies of transplantation immunity in hamsters. Ann. N.Y. Acad. Sci., 73: 676-687.
Craig, J. M. 1952 The histology of antigenically stimulated lymph nodes in rabbits given
ACTH or ACH (cortisone). Am. J. Path., 28:
629-652.
Craigmyle, M. B. 1958 Regional lymph node
changes induced by cartilage homo- and heterografts i n rabbits. J. Anat., 92: 74-83.
Foley, G. E., and A. H. Handler 1957 Differentiation of “normal” and neoplastic cells maintained i n tissue culture by implantation into
normal hamsters. Proc. SOC.Exp. Biol. Med.,
94: 661-664.
Lutz, B. R., G. P. Fulton, D. I. Patt and A. H.
Handler 1950 The growth rate of tumor
transplants i n the cheek pouch of the hamster
(Mesocricetus auratus.) Cancer Res., 10: 231232.
403
Lutz, B. R., G . P. Fulton, D. I. Patt, A. H. Handler
and D. F. Stevens 1951 The cheek pouch of
the hamster as a site for the transplantation of
a mbthylcholanthrene-induced sarcoma. Ibid.,
11: 64-66.
Ringertz, N., and C. A. Adamson 1950 The
lymph node response to various antigens. Acta
Path. Microbiol. Scand. Suppl., 86: 1-69.
Scothorne, R. J. 1956 The effect of cortisone
acetate on the response of the regional lymph
node to a skin homograft. J. Anat., 90: 117426.
Scothorne, R. J., and I. A. McGregor 1955 Cellular changes i n lymph nodes and spleen following skin homografting in the rabbit. Ibid., 89:
283-292.
Teilum, G . , H. C. Engback and M. Simonsen
1950 Effects of cortisone on plasma cells and
reticulo-endothelial system in hyperimmunized
rabbits. Acta Endocr., 5: 181-193.
Toolan, H. W. 1951 Successful subcutaneous
growth and transplantation of human tumors
in x-irradiated laboratory animals. Proc. SOC.
Exp. Biol. Med., 77: 572-578.
PLATE 1
EXPLANATION OF FIGURES
2
Hamster HT/28. The regional node which had been draining the cheek pouch with an
implanted amelanotic melanoma for 28 days. There is a distinct response in the size of
the cortex and in the presence of 26 highly active secondary centers. X 6.
3 Hamster HT/28. The non-regional node. Compare with the regional node seen in
figure 2. Smaller in size and fewer secondary centers.
4
Hamster HT/28. The cortex of the regional node. Note the large number of large
lymphocytes. X 430.
5 Hamster C8 /2 8 . The effects of cortisone acetate on a superficial cervical node. There
is an extreme reduction of the cortex and an increase in the number of cells within the
sinuses. x 8.
6 Hamster HT/28. The medulla of the regional node. Note the extreme plasma cell response. X 430.
404
LYMPH NODE RESPONSES TO TUMORS
David Shepro, Lester P. Eidelhoch and Donald I. Patt
PLATE 1
405